Protective immunity against human immunodeficiency virus-1 (HIV-1) is likely to require recognition of linear and conformation epitopes from multiple HIV antigens. Whether these responses can be elicited more effectively by virion-like structures or fused CTL epitopes is unknown.
The immune response to HIV infection in long-term non-progressors and HIV-exposed sex workers suggests that specific viral immunity may limit infection and the symptoms of disease. No single characteristic yet correlates with protective immunity, but studies in non-human of primates suggest that both humoral and cellular immunity are required for this response. Depletion of cytotoxic T cells (CTLs) in chronically-infected macaques enhances viremia. In humans, higher CTL responses correlate with lower viral load and stabilization of clinical symptoms. In animal models, passive transfer of neutralizing antibodies can also contribute to protection against virus challenge. Neutralizing antibody responses can also be developed in HIV-infected individuals and are associated with lower viral loads in long-term non-progressors. Though this neutralizing antibody response is uncommon, it is directed largely against the Env protein of the virus.
In early human vaccine trials, gp120 protein immunogens have yielded disappointing results: vaccine-induced antibodies have not been broadly neutralizing and have sometimes enhanced infection in vitro. Monomeric gp120 loses oligomer-dependent epitopes and does not include sequences in the ectodomain of the gp41 that become exposed during virus entry. It is assumed that broadly neutralizing antibodies bind to native gp120/gp41 complex on the surface of the virus rather than soluble gp120.
The development of a cytotoxic T lymphocyte (CTL) response to viruses is often crucial to the outcome of infections. Lysis of infected cells prior to the production of progeny virions may limit virus burst size (Yang, O et al., 1996, J. Virol., 70: 5799-5806), and HIV specific CD8+ cytotoxic T lymphocytes (CTL) have been shown to be important in viral clearance and in the control of initial HIV-1 spread (Borrow, P et al., 1994, J. Virol., 68: 6103-6110; Yang, O et al., 1996, J. Virol. 70: 5799-5806). CTL responses specific to HIV also contribute to reduction in viral load during acute and asymptomatic infection (Klein, M R, et al., 1995, J. Exp. Med. 181: 1365-1372; Moss, P A H et al., 1995, Proc. Natl. Acad. Sci. USA, 92: 5773-5777) and may be involved in protection against the establishment of persistent HIV infections (Rowland-Jones, S L et al., 1993, Lancet, 341: 860-861; Rowland-Jones, S L et. al., 1995, Nat. Med., 1: 59-64). High-frequency CTL responses to HIV-1 correlated with low viral load and slow disease progression in chronically infected individuals (Musey, L et al., 1997, N. Engl. J. Med., 337: 1267-1274; Ogg, G S et al., 1998, Science, 279: 2103-2106.). More compelling evidence of an antiviral effect of CD8+ cells was demonstrated in controlled studies in macaques, in which CD8+ cells were depleted in vivo using a monoclonal antibody. The viral loads in these animals increased or decreased as the CD8+ cells were depleted or reappeared, respectively (Jin, X et al., 1999, J. Exp. Med., 189: 991-998; Schmitz, J E, et al., 1999, Science, 283: 857-860). Therefore, induction of a CTL response specific to these proteins represents a desirable response in an HIV-1 vaccine.
HIV-1 internal structural and enzymatic proteins contain conserved domains that preserve their functions and thus exhibit less antigenic diversity that may elicit more effective CTL responses (Nixon, D F et al., 1988, Nature, 336: 484-487.). Efficient and durable CTL responses require endogenous antigen synthesis and processing. Current vaccine delivery techniques include immunization with live, attenuated viruses, inactivated recombinant virus infection (Letvin, N L, 1998, Science, 280: 1875-1880) or plasmid DNA expression vectors. A major obstacle in the induction of CTL responses with naked DNA or recombinant virus during development of an HIV vaccine is that the expression of HIV-1 structural and enzymatic genes is tightly regulated by the virus itself. The expression of these proteins is heavily dependent upon the existence of the Rev-responsive element (RRE) of HIV-1 in recombinant vectors (Cullen, B R, 1992, Microbiol. Rev., 56: 375-394; Felber, B K et al., 1989, Proc Natl Acad Sci USA, 86: 1495-1499). Poor expression is caused by the presence of AT rich inhibitory nucleotide sequences (INS) in the gag, pol and env genes, which inhibit the nuclear export and efficient expression of unspliced HIVI mRNAs. Early studies of DNA vaccination against HIV in mice required the inclusion of Rev in their expression vectors (Lu, S et al., 1995, Virology, 209: 147-154; Okuda, K et al., 1995, AIDS Res. Hum. Retroviruses, 11: 933-943; Wang, B et al., 1993, Proc. Natl. Acad. Sci. U.S. A 90: 4156-4160), but modification of INS has been shown to facilitate Rev-independent expression of HIV-1 Gag (Qiu, J-T et al., 1999, J. Virol., 73: 9145-9152; zur Megede, J et al., 2000, J Virol 74: 2628-2635), allowing detectable humoral and CTL responses against this protein (Qiu, J-T et al., 1999, J. Virol., 73: 9145-9152). These modified HIV-1 Gag genes produced viral-like particles of the expected density and morphology and induced an immune response to HIV-1 Gag after DNA immunization in mice (zur Megede, J et al., 2000, J Virol, 74: 2628-2635).